During an immune response against a microbial pathogen, nave T lymphocytes that recognize the microbe must give rise to two distinct types of progeny. Effector cells serve to provide acute host defense, whereas memory cells provide the ability to mount rapid recall responses should the host re-encounter the pathogen. Prior studies have demonstrated that a single nave CD8+ T lymphocyte is capable of generating all of the diverse cellular fates necessary for an immune response. The process by which a single activated T lymphocyte yields effector- and memory-fated progeny, and the timing at which these differentiation pathways begin to diverge, however, remains unanswered. We have recently shown that a T lymphocyte can undergo asymmetric division to give rise to two daughter cells that are differentially fated toward the effector and memory lineages. The overall goals of the active parent R01 grant are to investigate the functional consequences of asymmetric CD8+ T cell division and to elucidate the mechanisms by which asymmetric division leads to specification of the effector and memory T cell fates. In this revision application, we propose to develop novel single cell approaches to define the transcriptomic and epigenetic roadmap of an individual CD8+ T lymphocyte as it differentiates in vivo in response to a microbial pathogen. Accomplishment of the aims proposed herein is likely to yield important insights into the fundamental mechanisms by which asymmetric division influences cell fate specification and function, and may help to provide a framework for the rational development of vaccines.